EMDB-8607: Herpes Simplex Virion Primary Enveloped Virion

Basic information

• Entry: Database: EMDB / ID: EMD-8607
• Title: Herpes Simplex Virion Primary Enveloped Virion
• Map data: Masked HSV-PEV map

Sample

• Virus: Herpes simplex virus (type 1 / strain F)

Function / homology

Function:

chromosome organization => GO:0051276 / T=16 icosahedral viral capsid / deNEDDylase activity / viral genome packaging / viral tegument / viral capsid assembly / viral DNA genome replication / viral release from host cell / viral process / viral penetration into host nucleus / viral capsid / symbiont-mediated perturbation of host ubiquitin-like protein modification / ubiquitinyl hydrolase 1 / cysteine-type deubiquitinase activity / host cell cytoplasm / Hydrolases; Acting on peptide bonds (peptidases); Cysteine endopeptidases / symbiont entry into host cell / host cell nucleus / structural molecule activity / DNA binding

Domain/homology:

Herpesvirus large tegument protein deneddylase / Herpesvirus UL36 tegument protein / Herpesvirus UL35 / Herpesvirus UL35 family / Large tegument protein deneddylase / Herpesvirus tegument ubiquitin-specific protease (htUSP) domain profile. / Herpesvirus large tegument protein, USP domain / Herpesvirus tegument protein, N-terminal conserved region / Herpesvirus capsid vertex component 1 / Herpesvirus UL17 protein / Herpesvirus UL25 / Herpesvirus UL25 family / Herpesvirus capsid protein 2 / Herpesvirus capsid shell protein 1 / Herpesvirus VP23 like capsid protein / Herpesvirus capsid shell protein VP19C / Herpesvirus major capsid protein / Herpesvirus major capsid protein, upper domain superfamily / Herpes virus major capsid protein / Papain-like cysteine peptidase superfamily

Component:

Triplex capsid protein 1 / Capsid vertex component 1 / Triplex capsid protein 2 / Major capsid protein / Capsid vertex component 2 / Large tegument protein deneddylase / Small capsomere-interacting protein

• Biological species: Herpes simplex virus (type 1 / strain F)
• Method: single particle reconstruction / cryo EM / Resolution: 35.0 Å
• Authors: Fontana J / Newcomb WW / Winkler DC / Cheng N / Heymann JB / Steven AC
• Citation: Journal: mBio / Year: 2017; Title: The Primary Enveloped Virion of Herpes Simplex Virus 1: Its Role in Nuclear Egress.; Authors: William W Newcomb / Juan Fontana / Dennis C Winkler / Naiqian Cheng / J Bernard Heymann / Alasdair C Steven / ; Abstract: Many viruses migrate between different cellular compartments for successive stages of assembly. The HSV-1 capsid assembles in the nucleus and then transfers into the cytoplasm. First, the capsid buds through the inner nuclear membrane, becoming coated with nuclear egress complex (NEC) protein. This yields a primary enveloped virion (PEV) whose envelope fuses with the outer nuclear membrane, releasing the capsid into the cytoplasm. We investigated the associated molecular mechanisms by isolating PEVs from US3-null-infected cells and imaging them by cryo-electron microscopy and tomography. (pUS3 is a viral protein kinase in whose absence PEVs accumulate in the perinuclear space.) Unlike mature extracellular virions, PEVs have very few glycoprotein spikes. PEVs are ~20% smaller than mature virions, and the little space available between the capsid and the NEC layer suggests that most tegument proteins are acquired later in the egress pathway. Previous studies have proposed that NEC is organized as hexamers in honeycomb arrays in PEVs, but we find arrays of heptameric rings in extracts from US3-null-infected cells. In a PEV, NEC contacts the capsid predominantly via the pUL17/pUL25 complexes which are located close to the capsid vertices. Finally, the NEC layer dissociates from the capsid as it leaves the nucleus, possibly in response to pUS3-mediated phosphorylation. Overall, nuclear egress emerges as a process driven by a program of multiple weak interactions. On its maturation pathway, the newly formed HSV-1 nucleocapsid must traverse the nuclear envelope, while respecting the integrity of that barrier. Nucleocapsids (125 nm in diameter) are too large to pass through the nuclear pore complexes that conduct most nucleocytoplasmic traffic. It is now widely accepted that the process involves envelopment/de-envelopment of a key intermediate-the primary enveloped virion. In wild-type infections, PEVs are short-lived, which has impeded study. Using a mutant that accumulates PEVs in the perinuclear space, we were able to isolate PEVs in sufficient quantity for structural analysis by cryo-electron microscopy and tomography. The findings not only elucidate the maturation pathway of an important human pathogen but also have implications for cellular processes that involve the trafficking of large macromolecular complexes.

History

Deposition	Feb 19, 2017	-
Header (metadata) release	Mar 1, 2017	-
Map release	Jan 24, 2018	-
Update	Feb 14, 2018	-
Current status	Feb 14, 2018	Processing site: RCSB / Status: Released

Map

• File: Download / File: emd_8607.map.gz / Format: CCP4 / Size: 146 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
• Annotation: Masked HSV-PEV map
• Voxel size: X=Y=Z: 5.66 Å

Density

Contour Level	By EMDB: 200. / Movie #1: 200
Minimum - Maximum	-1032.047600000000102 - 724.766700000000014
Average (Standard dev.)	0.21723418 (±87.348039999999997)

• Symmetry: Space group: 1

Details

EMDB XML:

Map geometry	Axis order X Y Z Origin -168 -168 -168 Dimensions 337 337 337 Spacing 337 337 337
Cell	A=B=C: 1907.4199 Å α=β=γ: 90.0 °

CCP4 map header:

mode	Image stored as Reals
Å/pix. X/Y/Z	5.66	5.66	5.66
M x/y/z	337	337	337
origin x/y/z	0.000	0.000	0.000
length x/y/z	1907.420	1907.420	1907.420
α/β/γ	90.000	90.000	90.000
start NX/NY/NZ	0	0	0
NX/NY/NZ	28	11	56
MAP C/R/S	1	2	3
start NC/NR/NS	-168	-168	-168
NC/NR/NS	337	337	337
D min/max/mean	-1032.048	724.767	0.217

Supplemental data

Additional map: Non-masked HSV-PEV map

• File: emd_8607_additional.map
• Annotation: Non-masked HSV-PEV map

Sample components

Entire : Herpes simplex virus (type 1 / strain F)

• Entire: Name: Herpes simplex virus (type 1 / strain F)

Components

• Virus: Herpes simplex virus (type 1 / strain F)

Supramolecule #1: Herpes simplex virus (type 1 / strain F)

• Supramolecule: Name: Herpes simplex virus (type 1 / strain F) / type: virus / ID: 1 / Parent: 0 ; Details: US3 null mutant, a gift from Dr. Richard Roller, Dept. of Microbiology and Immunology, Carver College of Medicine, Univ. of Iowa; NCBI-ID: 10304 / Sci species name: Herpes simplex virus (type 1 / strain F) / Virus type: VIRION / Virus isolate: OTHER / Virus enveloped: Yes / Virus empty: No

Experimental details

Structure determination

• Method: cryo EM
• Processing: single particle reconstruction
• Aggregation state: particle

Sample preparation

• Buffer: pH: 7.5 / Details: PBS
• Vitrification: Cryogen name: ETHANE / Instrument: LEICA KF80

Electron microscopy

• Microscope: FEI/PHILIPS CM200FEG
• Electron beam: Acceleration voltage: 120 kV / Electron source: FIELD EMISSION GUN
• Electron optics: Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy
• Image recording: Film or detector model: GATAN ULTRASCAN 1000 (2k x 2k) / Average electron dose: 15.0 e/Ų

Image processing

• Particle selection: Number selected: 123 ; Details: PEV images of were picked manually, yielding a total of 123 particles.
• Startup model: Type of model: OTHER; Details: Origins and orientations were determined by projection matching, using as a starting model a map of B capsids obtained by in vitro maturation of purified procapsids (Aksyuk et al., 2015).
• Initial angle assignment: Type: RANDOM ASSIGNMENT
• Final angle assignment: Type: PROJECTION MATCHING
• Final reconstruction: Applied symmetry - Point group: I (icosahedral) / Resolution.type: BY AUTHOR / Resolution: 35.0 Å / Resolution method: FSC 0.33 CUT-OFF; Details: Reconstructions were calculated using breconstruct (from the Bsoft package), an algorithm that integrates images as central sections of Fourier space. An inverse Fourier transform was then calculated. Using an FSC cut-off of 0.3, the resolution of the reconstruction of C capsid-containing PEVs was estimated at ~3.5 nm for the entire particle and calculated (Cardone et al., 2013) at ~2 nm for the region around the capsid shell.; Number images used: 123
• Details: Images (2048 by 2048 pixels) were recorded using a CCD camera (Gatan) at a magnification of ~25,000x (giving a pixel size of 5.7 A) and processed using the Bsoft package (Heymann and Belnap, 2007) as previously described (McHugh et al., 2014).